Insights from entire mitochondrial genome sequences into the phylogeny of ticks of the genera Haemaphysalis and Archaeocroton with the elevation of the subgenus Alloceraea Schulze, 1919 back to the status of a genus.

We used entire mitochondrial (mt) genome sequences (14.5-15 kbp) to resolve the phylogeny of the four main lineages of the Haematobothrion ticks: Alloceraea, Archaeocroton, Bothriocroton and Haemaphysalis. In our phylogenetic trees, Alloceraea was the sister to Archaeocroton sphenodonti, a tick of an archetypal reptile, the tuatara, from New Zealand, to the exclusion of the rest of the species of Haemaphysalis. The mt genomes of all four of the Alloceraea species that have been sequenced so far had a substantial insert, 132-312 bp, between the tRNA-Glu (E) gene and the nad1 gene in their mt genomes. This insert was not found in any of the other eight subgenera of Haemaphysalis. The mt genomes of 13 species of Haemaphysalis from NCBI GenBank were added to the most recent data set on Haemaphysalis and its close relatives to help resolve the phylogeny of Haemaphysalis, including five new subgenera of Haemaphysalis not previously considered by other authors: Allophysalis (structurally primitive), Aboimisalis (structurally primitive), Herpetobia (structurally intermediate), Ornithophysalis (structurally advanced) and Segalia (structurally advanced). We elevated Alloceraea Schulze, 1919 to the status of genus because Alloceraea Schulze, 1919 is phylogenetically distinct from the other subgenera of Haemaphysalis. Moreover, we propose that the subgenus Allophysalis is the sister to the rest of the Haemaphysalis (14 subgenera) and that the 'structurally primitive' subgenera Hoogstraal and Kim comprise early diverging lineages. Our matrices of the pairwise genetic difference (percent) of mt genomes and partial 16S rRNA sequences indicated that the mt genome sequence of Al. kitaokai (gb# OM368280) may not be Al. kitaokai Hoogstraal, 1969 but rather another species of Alloceraea. In a similar way, the mt genome sequence of H. (Herpetobia) nepalensis Hoogstraal, 1962 (gb# NC_064124) was only 2% genetically different to that of H. (Allophysalis) tibetensis Hoogstraal, 1965 (gb# OM368293): this indicates to us that they are the same species. Alloceraea cretacea may be better placed in a genus other than Alloceraea Schulze, 1919. Reptiles may have been the host to the most recent common ancestor of Archaeocroton and Alloceraea.

Population structure and haplotype network analyses of Hyalomma anatolicum based on the large subunit ribosomal RNA (16S rRNA) gene.

Hyalomma anatolicum, an Anatolian hard tick is a well-recognized vector involved in the transmission of various pathogens to animals and humans. The present study elucidated the population structure and haplotype network of H. anatolicum based on the mitochondrial large subunit ribosomal RNA (16S rRNA) gene sequence. The population structure and haplotype network analysis of 75 sequences archived in the GenBank, including the 15 sequences generated herein, yielded 24 haplotypes. Haplotype 1 (Hap_1) was the predominant haplotype consisting of 45 sequences from India, China, Pakistan, Turkey, Egypt, Iraq, and Tajikistan. The complete haplotype network exhibited a stellate conformation, highlighting a recent population expansion. The overall dataset, together with the sequences corresponding to India, China, and Pakistan, showed a high haplotype (0.638 ± 0.065, 0.671 ± 0.103, 0.753 ± 0.099, and 0.854 ± 0.061, respectively) and low nucleotide (0.00407 ± 0.00090, 0.00525 ± 0.00196, 0.00680 ± 0.00233, and 0.00453 ± 0.00056, respectively) diversity, further emphasized a recent population expansion. The neutrality indices including Tajima's D, Fu and Li's D, and Fu and Li's F for the complete dataset (- 2.661, - 6.008, and - 5.649, respectively) as well as for the sequences from India (- 2.223, - 3.414, and - 3.567, respectively) were negative, suggesting deviation from neutrality and a recent population expansion. The present study provided novel insights into the population structure and haplotype networks of H. anatolicum based on the mitochondrial 16S rRNA gene, and the different tests inferred a low genetic differentiation and suggested a recent population expansion of this economically important tick species.

Application of DNA barcodes in the genetic diversity of hard ticks (Acari: Ixodidae) in Kazakhstan.

Forty-five tick species have been recorded in Kazakhstan. However, their genetic diversity and evolutionary relationships, particularly when compared to ticks in neighbouring countries, remain unclear. In the present study, 148 mitochondrial cytochrome c oxidase subunit I (COI) sequence data from our laboratory and NCBI (National Center for Biotechnology Information; https://www.ncbi.nlm.nih.gov/ ) data were used to address this knowledge gap. Phylogenetic analyses showed that i) Hyalomma anatolicum anatolicum (Koch, 1844) ticks from Jambyl Oblast (southeastern Kazakhstan) and Gansu Province (northwestern China) constituted a newly deviated clade; and ii) Dermacentor reticulatus (Fabricius, 1974) ticks from South Kazakhstan Oblast were closer to those in Romania and Turkey. The network diagram of haplotypes showed that i) the H-1 and H-2 haplotypes of Dermacentor marginatus (Sulzer, 1776) ticks from Zhetisu and Almaty were all newly evolved; and ii) the H-3 haplotypes of Haemaphysalis erinacei (Pavesi, 1884) from Almaty Oblast and Xinjiang Uygur Autonomous Region (northwestern China) were evolved from the H-1 haplotype from Italy. In the future, more COI data from different tick species, especially from Kazakhstan and neighbouring countries, should be employed in the field of tick DNA barcoding.

Initial study and phylogenetic analysis of hard ticks (Acari: Ixodidae) in Nantong, China along the route of avian migration.

The growing concern about migratory birds potentially spreading ticks due to global warming has become a significant issue. The city of Nantong in this study is situated along the East Asia-Australasian Flyway (EAAF), with numerous wetlands serving as roosting sites for migratory birds. We conducted an investigation of hard ticks and determined the phylogenetic characteristics of tick species in this city. We utilized three different genes for our study: the mitochondrial cytochrome oxidase subunit 1 (COX1) gene, the second internal transcribed spacer (ITS2), and the mitochondrial small subunit rRNA (12 S rRNA) gene. The predominant tick species were Haemaphysalis flava (H. flava) and Haemaphysalis longicornis (H. longicornis). Additionally, specimens of Haemaphysalis campanulata (H. campanulata) and Rhipicephalus sanguineus (R. sanguineus) were collected. The H. flava specimens in this study showed a close genetic relationship with those from inland provinces of China, as well as South Korea and Japan. Furthermore, samples of H. longicornis exhibited a close genetic relationship with those from South Korea, Japan, Australia, and the USA, as well as specific provinces in China. Furthermore, R. sanguineus specimens captured in Nantong showed genetic similarities with specimens from Egypt, Nigeria, and Argentina.

Redescription of adults and first description of the larva of Haemaphysalis (Rhipistoma) adleri Feldman-Muhsam, 1951 (Acari: Ixodidae), parasite of carnivorans in Western Asia.

The adults of Haemaphysalis (Rhipistoma) adleri Feldman-Muhsam, 1951 (Acari: Ixodidae) are redescribed and the larva of this species is described for the first time here. The adults of H. adleri that we studied were collected from various canid, felid and hyaenid carnivorans (Carnivora: Canidae, Felidae, Hyaenidae) as well as a hedgehog (Erinaceomorpha: Erinaceidae) in Iraq, Israel and West Bank. The males, females and larvae of H. adleri can be differentiated from Haemaphysalis (Rhipistoma) species occurring in the Palearctic portion of West Asia and Egypt as well those in the H. asiatica subgroup by the length of idiosomal setae, development and size of spurs on palpi, dental formula on the hypostome and size of spur on coxae. A lectotype of H. adleri has been designated and the geographic distribution and hosts of this tick species are discussed.

Some gastrointestinal nematodes and ixodid ticks shared by several wildlife species in the Kruger National Park, South Africa.

Parasite surveys were conducted for 1­2 years in the Kruger National Park (KNP), South Africa on blue wildebeest, impalas, greater kudus, common warthogs and scrub hares. The host associations of some of the gastrointestinal nematode species infecting ≥60% of at least one of the five host species, were determined. These were Agriostomum gorgonis, Cooperia acutispiculum, Cooperia connochaeti, Cooperia hungi, Cooperia neitzi, Cooperioides hamiltoni, Gaigeria pachyscelis, Haemonchus bedfordi, Haemonchus krugeri, Haemonchus vegliai, Impalaia tuberculata, Longistrongylus sabie, Strongyloides papillosus, Trichostrongylus deflexus and Trichostrongylus thomasi. Although the prevalence of Trichostrongylus falculatus did not exceed 50% in any host species, it was present in all five hosts. Nematodes in the KNP range from those exhibiting strict host associations to generalists. Nematode-host associations may be determined by host feeding patterns and habitat use. Eight ixodid tick species were commonly collected from the same animals and in 2­3 year long surveys from plains zebras and helmeted guinea fowls: Amblyomma hebraeum, Amblyomma marmoreum, Hyalomma truncatum, Rhipicephalus appendiculatus, Rhipicephalus decoloratus, Rhipicephalus evertsi evertsi, Rhipicephalus simus and Rhipicephalus zambeziensis. Host specificity was less pronounced in ixodid tick species than in nematodes and the immature stages of five tick species infested all host species examined.

Spatial clustering of fourteen tick species across districts of Zimbabwe.

BACKGROUND: Ticks transmit several diseases that result in high morbidity and mortality in livestock. Tick-borne diseases are an economic burden that negatively affect livestock production, cost countries billions of dollars through vaccine procurement and other disease management efforts. Thus, understanding the spatial distribution of tick hotspots is critical for identifying potential areas of high tick-borne disease transmission and setting up priority areas for targeted tick disease management. In this study, optimised hotspot analysis was applied to detect hotspots and coldspots of 14 common tick species in Zimbabwe. Data on the spatial distribution of tick species were obtained from the Epidemiology Unit of the Division of Veterinary Field Services of Zimbabwe. RESULTS: A total of 55,133 ticks were collected with Rhipicephalus decoloratus being the most common species (28.7%), followed by Amblyomma hebraeum (20.6%), and Rhipicephalus sanguineus sensu lato (0.06%) being the least common species. Results also showed that tick hotspots are species-specific with particular tick species occupying defined localities in the country. For instance, Amblyomma variegatum, Rhipicephalus appendiculatus, Rhipicephalus decoloratus, Rhipicephalus compostus, Rhipicephalus microplus, Rhipicephalus pravus, and Rhipicephalus simus were concentrated in the north and north eastern districts of the country. In contrast, Amblyomma hebraeum, Hyalomma rufipes, Hyalomma trancatum and Rhipicephalus evertsi evertsi were prevalent in the southern districts of Zimbabwe. CONCLUSION: The occurrence of broadly similar hotspots of several tick species in different districts suggests presence of spatial overlaps in the niche of the tick species. As ticks are vectors of several tick-borne diseases, there is high likelihood of multiple disease transmission in the same geographic region. This study is the first in Zimbabwe to demonstrate unique spatial patterns in the distribution of several tick species across the country. The results of this study provide an important opportunity for the development of spatially-targeted tick-borne disease management strategies.

Ticks (Acari: Ixodoidea) associated with mammals in Colombia: a historical review, molecular species confirmation, and establishment of new relationships.

Ticks are considered the second most important vectors of pathogens worldwide, after mosquitoes. This study provides a systematic review of vector-host relationships between ticks and mammals (domestic and wild) and consolidates information from studies conducted in Colombia between 1911 and 2020. Using the PRISMA method, 71 scientific articles containing records for 51 tick species (Argasidae and Ixodidae) associated with mammals are reported. The existing information on tick-mammal associations in Colombia is scarce, fragmented, or very old. Moreover, 213 specimens were assessed based on morphological and molecular analyses, which allowed confirming eight tick species associated with mammals: Amblyomma calcaratum, Amblyomma dissimile, Amblyomma mixtum, Amblyomma nodosum, Amblyomma ovale, Amblyomma varium, Ixodes luciae, and Ixodes tropicalis. Several tick species are molecularly confirmed for Colombia and nine new relationships between ticks and mammals are reported. This research compiles and confirms important records of tick-mammal associations in Colombia.

Ticks and tick-borne pathogens infecting livestock and dogs in Tchicala-Tcholoanga, Huambo Province, Angola.

The diversity of ticks and tick-borne pathogens (TBPs) infesting domestic animals in Tchicala-Tcholoanga, Angola, in 2016 was investigated. Seventeen tick species were recorded, Amblyomma pomposum being the most abundant on cattle (40%), goats (38%) and sheep (35%); Rhipicephalus turanicus was the most abundant on dogs (46%). This study presents new records of Haemaphysalis paraleachi, R. compositus, R. kochi and R. sulcatus in Angola, the first georeferenced population of Ha. leachi in southern Africa and the second record of R. microplus in Angola. Using the reverse line blot (RLB) hybridisation assay, fifteen TBP species were detected in blood samples from cattle (n = 88), goats (n = 82), sheep (n = 85) and dogs (n = 85). F The most frequently detected species were Theileria velifera in cattle (78%), Theileria ovis in sheep (80%) and Babesia vogeli in dogs (35%). Species-specific quantitative PCR assays detected Babesia bigemina in 43% (35/80) of blood samples of cattle, while E. ruminantium was detected in 4% (3/70) of blood samples and in 7% of A. pomposum ticks. Anaplasma platys was detected from cattle (18%) and sheep (6%) during RLB analysis. These findings constitute pioneering research in Angola.

Potential tick vectors for Theileria equi in Israel.

Theileria equi Mehlhorn and Schein, 1998 (Piroplasmida: Babesiidae) is an important tick-borne pathogen of horses that is highly endemic in many parts of the world, including Israel. The present study evaluated the potential roles of five hard tick species [Hyalomma excavatum Koch, 1844; Hyalomma marginatum Koch, 1844; Rhipicephalus turanicus Pomerantsev 1936; Rhipicephalus annulatus Say, 1821; Haemaphysalis parva (Neumann, 1897) (all: Ixodida: Ixodidae)], previously found to infest horses in Israel, in acting as vectors for piroplasmosis. For this, DNA was extracted from whole ticks and, when possible, from the salivary glands in each species (n = 10-59). Polymerase chain reaction amplification and sequencing of the 18S rRNA gene were used to detect T. equi in 48 of the 127 ticks (37.8%) and in 21 of the 90 extracted salivary glands (23.3%) in all five species. All but two sequences were classified as T. equi genotype A; the remaining two were classified as genotype D. The findings of this study point to Ha. parva and R. annulatus as potential novel vectors of T. equi, and suggest that parasite genotype selection occurs within the tick vector.

Molecular cloning, expression and impact of ribosomal protein S-27 silencing in Haemaphysalis longicornis (Acari: Ixodidae).

Ticks, obligatory blood-feeding arthropods, are a major pathogen vector in humans and animals worldwide. Anti-tick vaccines are an exciting alternative to chemical acaricides for controlling these disease-transmitting vectors. However, identification of protective antigens for anti-tick vaccine development is challenging. Different ribosomal proteins play multifunctional roles in tick survival and feeding. Here, we first report the cloning and molecular characterization of ribosomal protein S27 (RPS-27) from the hard tick Haemaphysalis longicornis. We identified a complete open reading frame (ORF) of RPS-27: a 255-bp (base pair) cDNA encoding a mature protein of 84 amino-acid residues with a 9.4-kDa predicted molecular mass. Amino-acid sequence analysis revealed that RPS-27 was highly conserved among different tick and vertebrate animals with identity ranges of 97-98% and 60-85%, respectively. Phylogenetic tree analysis showed that RPS-27 from different tick species clustered together. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that the RPS-27 mRNA transcript was expressed in all life stages. At the tissue level, it was more highly expressed in the salivary gland than in the midgut for both the fed and unfed conditions, which indicates a role for RPS-27 in tick feeding. In vitro analysis showed that recombinant RPS-27 (10-RPS-27) was successfully expressed in a pGEMEX-2 vector with an estimated 45-kDa molecular mass. The functional importance of RPS-27 was determined by gene silencing through RNA interference (RNAi). RPS-27 silencing showed a significant (P < 0.05) reduction of feeding abilityand engorgement weight after the blood meal in both nymph and adult female ticks and also significantly (P < 0.05) reduced molting rate in nymph. In addition, RPS-27 silencing in eggs led to abnormalities in shape and hatching. Taken together, our results suggest that RPS-27 is an important molecule that plays multiple roles in the tick life cycle including in both feeding and reproduction. Therefore, RPS-27 is an exciting target for future tick control strategies.

Morphological and molecular identification of ixodid tick species (Acari: Ixodidae) infesting cattle in Uganda.

In Uganda, the role of ticks in zoonotic disease transmission is not well described, partly, due to limited available information on tick diversity. This study aimed to identify the tick species that infest cattle. Between September and November 2017, ticks (n = 4362) were collected from 5 districts across Uganda (Kasese, Hoima, Gulu, Soroti, and Moroto) and identified morphologically at Uganda Virus Research Institute. Morphological and genetic validation was performed in Germany on representative identified specimens and on all unidentified ticks. Ticks were belonging to 15 species: 8 Rhipicephalus species (Rhipicephalus appendiculatus, Rhipicephalus evertsi evertsi, Rhipicephalus microplus, Rhipicephalus decoloratus, Rhipicephalus afranicus, Rhipicephalus pulchellus, Rhipicephalus simus, and Rhipicephalus sanguineus tropical lineage); 5 Amblyomma species (Amblyomma lepidum, Amblyomma variegatum, Amblyomma cohaerens, Amblyomma gemma, and Amblyomma paulopunctatum); and 2 Hyalomma species (Hyalomma rufipes and Hyalomma truncatum). The most common species were R. appendiculatus (51.8%), A. lepidum (21.0%), A. variegatum (14.3%), R. evertsi evertsi (8.2%), and R. decoloratus (2.4%). R. afranicus is a new species recently described in South Africa and we report its presence in Uganda for the first time. The sequences of R. afranicus were 2.4% divergent from those obtained in Southern Africa. We confirm the presence of the invasive R. microplus in two districts (Soroti and Gulu). Species diversity was highest in Moroto district (p = 0.004) and geographical predominance by specific ticks was observed (p = 0.001). The study expands the knowledge on tick fauna in Uganda and demonstrates that multiple tick species with potential to transmit several tick-borne diseases including zoonotic pathogens are infesting cattle.

Checklist of the hard tick (Acari: Ixodidae) fauna of Hungary with emphasis on host-associations and the emergence of Rhipicephalus sanguineus.

Hungary is situated in the southern part of Central Europe, next to the northern boundary of the Mediterranean region. This geographical position may allow the northward expansion of Mediterranean ixodid tick species into Hungary, particularly in the era of warming climate. During the past 14 years numerous surveys have been published on the species and activity of hard ticks occurring in the country. However, it was 60 years ago that the last comprehensive review of ixodid ticks of Hungary was published, and only in Hungarian language. The purpose of the present checklist is to provide a comprehensive and complete overview of the ixodid fauna of Hungary, based on tick reports published so far in Hungarian or English, also including hitherto unpublished data. Altogether 27 hard tick species were identified in Hungary, of which 21 can be regarded as indigenous. Most importantly, the autochthonous occurrence of Rhipicephalus sanguineus sensu lato was not known prior to 2005, but during the last 14 years increasing numbers of cases have been reported, attesting the emergence of this tick species in Hungary. Whereas R. sanguineus sensu lato was always associated with dogs and cats in Hungary, other tick species show differences in host associations according to habitat type, seasonal activity and questing height. Changes in the distribution, abundance and seasonality of a few tick species were also noted.

Description of a new species of Haemaphysalis Koch, 1844 (Acari: Ixodidae) from the H. (Rhipistoma) asiatica subgroup, parasite of an endemic Malagasy carnivoran (Carnivora: Eupleridae).

Haemaphysalis (Rhipistoma) galidiae n. sp. (Acari: Ixodidae) is described herein based on males and a single female. Adults of H. galidiae n. sp. were collected from the ring-tailed vontsira, Galidia elegans Geoffroy Saint-Hilaire (Carnivora: Eupleridae) on Madagascar. Males and the single female of this new species morphologically are most close to H. obtusa Dönitz, 1910, known from various Malagasy euplerid carnivorans, including G. elegans, as well as introduced carnivorans, but can be differentiated from it by the shape of the female genital aperture, shape of posterolateral margin of palpal segment II ventrally in both male and female and shape and size of spurs on palpal segment II, coxa I and trochanter I in both sexes.

Description of two new species of Haemaphysalis Koch, 1844 (Acari: Ixodidae) from the H. (Rhipistoma) spinulosa subgroup, parasites of carnivores and other mammals in Africa.

Haemaphysalis (Rhipistoma) latitudinis n. sp. (Acari: Ixodidae) and H. (R.) setosa n. sp. are described based on adults. Adults of H. (R.) latitudinis n. sp. were mostly collected from carnivores (Carnivora: Canidae, Felidae, Herpestidae, Mustelidae, Viverridae), few specimens were found on rodents (Rodentia: Muridae), bat (Chiroptera: Molossidae) and domestic cattle (Artiodactyla: Bovidae) in Benin, Central African Republic, Côte d'Ivoire, Democratic Republic of the Congo, Ethiopia, Ghana, Kenya, Nigeria, South Sudan and Uganda. Adults of H. (R.) setosa n. sp. were mostly found on carnivores (Carnivora: Felidae, Herpestidae, Viverridae), few specimens were found on rodents (Rodentia: Muridae), hare (Lagomorpha: Leporidae), hyrax (Hyracoidea: Procaviidae) and human (Primates: Hominidae) in Kenya. Males and females of both species can be differentiated from each other and other H. spinulosa-like ticks by their size, pattern of punctations on conscutum/scutum, size of setae, shape of genital structures, size and shape of spiracular plates, dorsal cornua, posterodorsal and posteroventral spurs on palpal segment II and spur on coxae and dentition on hypostome.

Early Questing by Lone Star Tick Larvae, New York and Massachusetts, USA, 2018.

Subtropical lone star tick larvae typically emerge in late summer. We found clusters of host-seeking lone star tick larvae during early June 2018 in New York and Massachusetts, USA. Invasion and persistence of this tick in more northern locations may have been promoted by adaptation to an accelerated life cycle.

Long term persistence of introduced Amblyomma geoemydae tick population under indoor conditions in Austria.

An indoor terrarium population of Amblyomma geoemydae was established subsequent to the import of a single yellow-marginated box turtle Cuora flavomarginata. This indoor tick population revealed an unexpected resistance against de-ticking trials, with persistence between 2010 and 2015, when the ticks were successfully eliminated. Ticks were collected from the bodies and shells of turtles, as well as from terraria soil. Species diagnosis of ticks was carried out according to distinguishable morphological characters and supported by molecular analysis using DNA-barcoding. Introduced exotic ticks are potential vectors of pathogens and can have an impact on wildlife, domestic animals and the human population. This case emphasizes the need for sharp surveillance and control measures on imported reptiles.

Identifying the tick Amblyomma javanense (Acari: Ixodidae) from Chinese pangolin: generating species barcode, phylogenetic status and its implication in wildlife forensics.

Zoonotic diseases transmitted through ticks and other ectoparasites often travel across the globe with illegally traded wildlife parts and products. In this study, we analyzed a confiscated case of pangolin scales and observed a few dead ticks attached. On genetic analysis, the pangolin scales were identified to be originated from Chinese pangolin (Manis pentadactyla), an IUCN listed Critically Endangered species, and ticks were identified as Amblyomma javanense. Here, we provide the first authentic physical record of A. javanense from India as a parasite of Chinese pangolin and also generated its species DNA barcode that may be useful for biologists working on ticks in species validation and constructing phylogenies across the globe.

Molecular detection of spotted fever group rickettsiae in ticks parasitizing pet dogs in Shihezi City, northwestern China.

A total of 178 adult ticks were collected from 32 pet dogs from five veterinary clinics in Shihezi City, Xinjiang Uygur Autonomous Region (XUAR), northwestern China. All the ticks were identified by comprehensive morphological and genetic analyses, and rickettsiae were detected by seven Rickettsia-specific genetic markers in the ticks. The ticks collected were identified as Rhipicephalus sanguineus sensu lato. Twenty-one of the 178 samples (11.8%) were positive for rickettsiae. Among these, in 13 (61.9%) samples Candidatus R. barbariae were identified, in five (23.8%) samples R. massiliae, and in three (14.3%) samples R. conorii. This study indicates that more attention should be paid to rickettsial infection in pet dogs and their ticks, because the latter may pose an epidemiological risk for tick-borne transmission of rickettsiae to human beings.